Coordination of the cytoskeletons of neighboring cells is an important component of morphogenesis during animal development. As a model system to study this phenomena I am using the epidermis of Drosophila. This tissue elaborates a large number of polarized structures such as sensory bristles and hairs. In any body region these are typically aligned in parallel giving the tissue a distinct polarity. For example, the wing is covered with an array of distally pointing hairs that are formed in the pupae from F actin filled microvillus-like prehairs. Mutations in tissue polarity genes disrupt the normal hair polarity pattern. We have shown that the tissue polarity genes comprise a genetic pathway that regulates hair polarity via Controlling the subcellular location for prehair development. The focus of this proposal is the further dissection of this pathway. The frizzled (fz) tissue polarity gene encodes a transmembrane protein that is essential for both the transmission and transduction of an intercellular polarity signal. The sequencing of mutations isolated in vivo suggests that distinct domains of the Fz protein are involved in each of these functions. We will use in vitro mutagenesis and germ line transformation to test this hypothesis. The inturned (in) gene is downstream of fz and functions as an inhibitor of prehair initiation. We have recently cloned and sequenced this gene. We propose to use immunostaining to determine its subcellular localization, germ line transformation to determine if overexpression results in a gain of function mutation, and in vitro mutagenesis and germ line transformation to test if a presumptive extracellular RGD sequence causes the In protein to function as a ligand for an integrin. We have preliminary data that indicates that the microtubule cytoskeleton is essential for the normal regulation of prehair initiation. We will confirm this observation and attempt to determine the mechanism involved. Finally we will conduct a large mutant screen to identify additional tissue polarity genes. Via genetic interaction experiments we will attempt to place these new genes into the existing pathway model.